Available applications

Background of ATF3 antibody

This protein binds the cAMP response element (CRE) (consensus: 5\'-GTGACGT[AC][AG]-3\'), a sequence present in many viral and cellular promoters. ATF3 represses transcription from promoters with ATF sites. It may repress transcription by stabilizing the binding of inhibitory cofactors at the promoter. Isoform 2 activates transcription presumably by sequestering inhibitory cofactors away from the promoters.Activating transcription factor 3 is a member of the mammalian activation transcription factor/cAMP responsive element-binding (CREB) protein family of transcription factors. Multiple transcript variants encoding two different isoforms have been found for this gene. The longer isoform represses rather than activates transcription from promoters with ATF binding elements. The shorter isoform (deltaZip2) lacks the leucine zipper protein-dimerization motif and does not bind to DNA, and it stimulates transcription presumably by sequestering inhibitory co-factors away from the promoter. It is possible that alternative splicing of the ATF3 gene may be physiologically important in the regulation of target genes.

Western blot of E.coli whole cell extract transfected with GST epitope tagged human ATF3. Anti-ATF3 detects a band ~48 kDa corresponding to recombinant human ATF3. Anti-GST epitope tag antibodyconfirms the composition of the recombinant band (not shown). The protein was transferred to nitrocellulose using standard methods. Afterblocking with 5% goat serum and 0.5% non fat milk in PBS, the membrane was probed with the primary antibody diluted 1:200 in 0.2X blocking buffer in PBS overnight at 4°C. Reaction was followed bywashes and reaction with a 1:5000 dilution of IRDye(TM)800 conjugated Gta-Rabbit IgG [H&L] for 30 min at room temperature. LICOR's Odyssey® Infrared Imaging System was used to scan and process the image. Other detection systems will yield similar results.

Western blot of mammalian whole cell extract transfected with HA epitope tagged human ATF3. Anti-ATF3 detects a band ~31 kDa corresponding to recombinant human ATF3. Immunostaining using anti-HA epitope tag antibody confirms the composition of the recombinant band (not shown). The protein was transferred to nitrocellulose in 30 minutes using standard methods. After blocking with 5% goat serum and 0.5% non-fat milk in PBS, the membrane was probed with the primary antibody diluted 1:200 in 0.2X blocking buffer in PBS overnight at 4°C. Reaction was followed by washes and reaction with a 1:5000 dilution of IRDye(TM)800 conjugated Gt-a-Rabbit IgG [H&L] for 30 min at room temperature. LICOR's Odyssey® Infrared Imaging System was used to scan and process the image. Other detection systems will yield similar results.

Western blot of mammalian whole cell extract transfected with HA epitope tagged human ATF3. µgeneTex's Affinity purified anti-ATF3 detects a band ~31 kDa corresponding to recombinant human ATF3. Immunostaining using GeneTex's anti-HA epitope tag antibody confirms the composition of the recombinant band (not shown). The protein was transferred to nitrocellulose in 30 minutes using standard methods. After blocking with 5% goat serum and 0.5% non-fat milk in PBS, the membrane was probed with the primary antibody diluted 1:200 in 0.2X blocking buffer in PBS overnight at 4°C. Reaction was followed by washes and reaction with a 1:5000 dilution of IRDye800 conjugated goat anti-Rabbit IgG [H&L]) for 30 min at room temperature. LICOR's Odyssey® Infrared Imaging System was used to scan and process the image. Other detection systems will yield similar results.

Western blot of E.coli whole cell extract transfected with GST epitope tagged human ATF3. µgeneTex's Affinity purified anti-ATF3 detects a band ~48 kDa corresponding to recombinant human ATF3. Immunostaining using GeneTex's anti-GST epitope tag antibody confirms the composition of the recombinant band (not shown). The protein was transferred to nitrocellulose using standard methods. After blocking with 5% goat serum and 0.5% non fat milk in PBS, the membrane was probed with the primary antibody diluted 1:200 in 0.2X blocking buffer in PBS overnight at 4°C. Reaction was followed by washes and reaction with a 1:5000 dilution of IRDye800 conjugated goat anti-Rabbit IgG [H&L]) for 30 min at room temperature. LICOR's Odyssey® Infrared Imaging System was used to scan and process the image. Other detection systems will yield similar results.

Immunohistochemical analysis of ATF3 staining in human lung cancer formalin fixed paraffin embedded tissue section. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0). The section was then incubated with the antibody at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.

HEK293T cells were transfected with the pCMV6-ENTRY control (Left lane) or pCMV6-ENTRY ATF3 (RC202897, Right lane) cDNA for 48 hrs and lysed. Equivalent amounts of cell lysates (5 ug per lane) were separated by SDS-PAGE and immunoblotted with anti-ATF3.(1:2000)